In aging, decreased levels of circulating estrogen are associated with memory impairment and cognitive decline, but the molecular events underlying this dysfunction are poorly understood. Therefore, the broad, long-term goal is to understand how estrogen supports synaptic activity, with particular focus on the hippocampus, the area of the brain associated with learning and memory. The specific goal of this project is to define the molecular pathways by which estroqen regulates NR1, the obligate subunit of the NmethyI- D-aspartate (NMDA) receptor (NMDAR). Mice lacking NR1 display an absence of hippocampal synaptic plasticity and have impaired spatial memory. In wild-type animals, endogenous increases in estrogen across the estrous cycle are associated with improved spatial memory, increased NR1 protein levels, and increased NMDAR binding in the hippocampus. Preliminary data indicate that estrogen also can directly increase NR1 mRNA levels in vitro, yet there are no estrogen receptor (ER) response elements in the genomic NR1 promoter. Therefore, the testable hypothesis is that estrogen stimulates NR1 activity and thus supports consequent synaptic function -- using non-genomic mechanisms on both promoter activation as well as post translational modification. The project has four independent yet inter-related aims: Aim 1 will quantify estrogen-stimulated increases in NR1 mRNA levels by using organotypic hippocampal slice cultures and quantitative RT-PCR to achieve significant sensitivity over current in vivo approaches. Aim 2 will quantify the non-genomic actions of estrogen on the NR1 promoter using NR1 reporter constructs and mutant variants. Aim 3 will determine how different ER isoforms (ERalpha or ERbeta) lead to NR1 promoter modulation. Aim 4 will determine how non-genomic estrogen actions post-translationally phosphorylate NR1 to modify NMDAR localization, sensitivity, and activity at the synapse. The NMDAR is the major glutamatergic receptor in the hippocampus, and deficit in NR1 is associated with aging and human neurologic diseases. This project will elucidate how estrogen nongenomically stimulates and supports cognitive function by promoting synaptic activity in hippocampal neurons, and, by corollary, will explain one way the loss of estrogen may affect neuronal dysfunction observed in the aging process.